tbuckley/cabot-search huggingface dataset:
Abstract P1157: Perinuclear Damage From Nuclear Envelope Deterioration Elicits Stress Responses That Contribute To <i>Lmna</i> Cardiomyopathy
Authors: Kunal Sikder, Elizabeth Phillips, Jasmine Saunders, David Mothy, Jason C Choi
Journal: Circulation Research (2023)

Mutations in the LMNA gene encoding nuclear lamins A/C cause a diverse array of tissue-selective diseases, with the heart being the most commonly affected organ. Despite progress in understanding the molecular perturbations emanating from LMNA mutations, an integrative understanding of the pathogenesis leading to cardiac dysfunction remains elusive. Nuclear envelope damage resulting from LMNA mutations is widely observed and accepted to be involved in the disease pathogenesis but how it contributes is not well understood. To this end, we performed a detailed kinetic study using a temporally controlled, cardiomyocyte-specific deletion model of Lmna to elucidate the progression of the disease. We found that cardiomyocyte-specific Lmna deletion in 12 weeks-old adult mice led to rapid cardiomyopathy with pathological remodeling. Prior to the onset of cardiac dysfunction, lamin A/C-depleted cardiomyocytes displayed nuclear envelope deterioration and extensive damage to the perinuclear space. Perinuclear mitochondria and golgi are particularly affected, leading to mitochondrial loss as well as golgi dilation/fragmentation and CREB3-mediated golgi stress activation. Translatome profiling identified upregulation of Med25, a transcriptional co-factor that can selectively dampen UPR axes. Autophagy is disrupted in the hearts of these mice, which can be recapitulated by disrupting the golgi or inducing nuclear damage by increased matrix stiffness. Systemic administration of pharmacological modulators of autophagy or ER stress significantly improved the cardiac function. These studies support a hypothesis wherein stress responses emanating from the perinuclear space contribute to the development of LMNA cardiomyopathy.
Abstract P1141: Molecular Insights Into The Phenotypic Heterogeneity Observed In Two Different Murine Models Of Hypertrophic Cardiomyopathy (HCM)
Authors: Tilo Thottakara, Carlin S Long, Arun Padmanabhan, Jeffrey Olgin, M Roselle Abraham
Journal: Circulation Research (2023)

Background: Phenotypic variability in HCM has long been appreciated to correlate with genotype. However, the molecular drivers underpinning these differences remain poorly understood. We hypothesized that the variable HCM phenotypes observed in 2 different widely used HCM mouse models (R403Q-MyHC, R92W-TnT) result from complex interactions between a variety of cardiac cell types. We performed unbiased assessments of gene expression and chromatin accessibility in mutant and littermate control hearts at an early disease stage, when MyHC-mutants have higher LV-mass and normal LV-function, while TnT-mutants have similar LV-mass, hypercontractility and diastolic dysfunction, compared to littermate-controls. Methods: Nuclei were isolated from hearts of 5-week-old mutant and control animals and subjected to the standard linked ATAC and gene expression profiling pipeline from 10X Genomics w/o enrichment/sorting. Clustering, differential gene expression and transcription factor (TF) motif enrichment was performed using Seurat, ArchR; SCENIC was used to predict TF (Regulon)-activation from gene expression data and Cellchat for cellular crosstalk. Results: Integrated single-nuclei data-based clustering identified all cardiac cell types. Both mutant ventricular(V)-myocytes had similar dysregulation (80-90% agreement) of gene expression and TF enrichment, with matching of 23/30 top TF-motifs, and agreement with gene expression-based TF-prediction. Both mutant V-myocytes had activation of SWI/SNF complex TFs (Jun, Fos, Smarc family), however activation of SREBF-1/2, Atf5, Creb3l2, Egr2 (drivers of fibroblast activation), and Klf2, Klf4, Bcl6b (drivers of shear stress-mediated gene expression in endothelial cells (ECs)) was only present in TnT-mutants. Activation of VCAM signaling from V-myocytes-to-ECs, and PDGF signaling to VSMCs is predicted in both mutants, but TGF-β signaling to fibroblasts, endothelin-1 signaling to V-myocytes, was only predicted in TnT-mutants. Conclusions: Mutant sarcomeric proteins activate the SWI/SNF complex that drives V-myocyte hypertrophy at early disease stage. Mutation-specific activation of TF-programs in non-myocyte cells are important drivers of cardiac phenotypic variability in HCM.
Abstract P1137: Novel Transcriptomic Interactions In Human Cardiomyopathic Heart Failure
Authors: Gina Qualter, Viswanathan Rajagopalan, Kenneth S Campbell
Journal: Circulation Research (2023)

Noncoding regions span ~98% of the human genome and are emerging as key players in cardiovascular pathologies. However, the role of human noncoding RNAs and their interactions with other RNAs and protein-coding genes in ischemic (ICM) and nonischemic cardiomyopathic (NICM) heart failure is not clear. Twenty-eight patients (53±11 years; mix of gender and race) were studied using Tukey’s analysis of variance and post-hoc analyses. Significantly increased left ventricular (LV) internal diameter (diastole and systole; p&lt;0.01) and decreased ejection fraction (EF:p&lt;1E-10) were observed in both ICM (EF:17.5%±3.5) and NICM (EF:20.6%±3.2) compared to organ donor controls (EF:56.6%±6). End-systolic volume was increased in ICM, and QTc and QRS duration were prolonged in NICM (p&lt;0.05). Following blinded quality control validation of both LV RNA and data, blinded whole transcriptomics (n=4/group) revealed differential expression of numerous long noncoding (lnc)RNAs, micro(mi)RNAs, and mRNAs (p-corrected/adjusted; padj&lt;0.05). We uncovered 251 novel lncRNAs, 137 novel mature miRNAs, and 286 novel mRNAs (padj&lt;0.05). Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), Reactome and Disease Ontology enrichment analyses revealed several significant (padj&lt;0.05) pathways in cancer, neuro/macular degeneration, neuroembryological development, endocrine, inflammation, bone remodeling, vein disease, etc. Seven lncRNA-associated protein-protein interaction networks were identified in ICM (&gt;700 confidence score). Co-expression analysis revealed highly significant lncRNA-mRNA interactions with Pearson Correlation Coefficient of &lt;0.95 and &gt;0.95 with p&lt;0.00001. GO/KEGG also revealed miRNA-mRNA interactions related to bacterial/viral, parathyroid, cancer, mTOR, etc. (padj&lt;0.05). Importantly, several genes and interactions in all the above RNA species and multiple enrichment pathways distinguished ICM versus NICM (padj&lt;0.05). We also observed several lncRNA alternative splicing events (padj&lt;0.0004; false discovery rate&lt;0.05), numerous missense and silent mutations, and minimal nonsense mutations. Together, this study offers novel transcriptomic and interactomic understanding in human ICM and NICM heart failure.
Abstract P1174: Nicotine Alters PHLPP Isoforms In The Heart To Increase Susceptibility To Injury
Authors: Khaja Shameem Mohammed Abdul, Alyssa Guerrero, Amogh Kulkarni, Tan Nguyen, Nicole H Purcell
Journal: Circulation Research (2023)

Rationale: The impact of nicotine exposure through vaping on cardiovascular (CV) protective signaling basally or following injury remains largely unknown. Recent studies have shown that PHLPP1 deletion increased activity of AKT in the heart and reduced injury whereas PHLPP2 deletion exacerbated cardiac injury. Preliminary studies in vivo found that systemic exposure to nicotine increased PHLPP1 and decreased PHLPP2 expression in heart. Determining the mechanism of nicotine induced PHLPP alterations will demonstrate the impact of nicotine use on CV function and physiology and whether targeting PHLPP will reduce injury. Hypothesis: We hypothesize that nicotine compromises metabolic adaptation of the heart to injury by altering PHLPP isoforms and dysregulating mitochondrial biogenesis and oxidative stress signaling. Methods: Male and female wild-type (WT, n=4) and PHLPP1 knockout (KO, n=4) 4-6 weeks of age were implanted with osmotic minipumps containing nicotine (5mg/kg/day) or saline for 7 days. For in vitro study, NRVMs were isolated and PHLPP isoforms were targeted with siRNA or overexpressed using adenoviruses. Cardiomyocytes were stimulated with nicotine (100nM, 48hrs). Results: Nicotine increased PHLPP1 expression (1.7 ± 0.25-fold vs. WT saline) in the heart and decreased PHLPP2 (0.8 ± 0.04-fold vs. WT saline) whereas the KO exhibited increased PHLPP2 (1.5 ± 0.04-fold vs KO saline) levels. Nicotine administration significantly decreased genes involved in mitochondrial biogenesis and antioxidant signaling in WT mice while significantly increasing in KO hearts. Cardiomyocytes stimulated with nicotine (48h) significantly increased PHLPP1 (1.9 ± 0.39-fold vs PBS) and ROS. Nicotine significantly decreased mitochondrial respiration as did overexpression of PHLPP1. Finally, knockdown of PHLPP1 significantly increased mitochondrial respiration. Conclusions: Increases in PHLPP1 in the heart and cardiomyocytes from nicotine exposure has detrimental effects on oxidative stress and mitochondrial respiration. Future studies will investigate the molecular mechanisms by which nicotine alters PHLPP protein expression in heart and whether targeting PHLPP1 could reduce cardiovascular injury induced by nicotine exposure.
Abstract P2021: Targeting Of Cdc42-interacting Protein 4 -calcineurin Signaling Complexes Is Cardioprotective During Ischemia/Reperfusion Injury
Authors: Anne-Maj Samuelsson, Yang Li, Jinliang Li, Michael S Kapiloff
Journal: Circulation Research (2023)

Myocardial infarction (MI) is a major cause of mortality and morbidity worldwide. Heart failure due to MI results from both acute ischemic/reperfusion (I/R) injury and subsequent post-MI pathological cardiac remodeling. The Ca 2+ /calmodulin-dependent protein phosphatase calcineurin (CaN), which promotes pathological cardiac remodeling, has been problematic as a therapeutic target due to its role in cardioprotection, including during I/R injury. We have shown that a select pool of CaNAβ holoenzyme is localized within the cardiomyocyte by scaffold protein Cdc42-Interacting Protein 4 (CIP4/TRIP10), such that the CaNAβ N-terminal polyproline (PP) domain binds directly to the CIP4 C-terminal SH3 domain. Pathological cardiac remodeling induced by pressure overload was inhibited by CIP4 gene knock-out and by inhibition of CaNAβ binding to CIP4 using an adeno-associated virus (AAV) vector that expresses the CaNAβ PP peptide. We now consider whether targeting CIP4-CaNAβ signalosomes is beneficial in ischemic cardiomyopathy. To test the relevance of CIP4 in vivo to ischemic reperfusion (IR) injury, tamoxifen-inducible, cardiac-specific CIP4 knock-out (CIP4 CKO), control Tg(Myh6-cre/Esr1*), and control CIP4 f/f mice were subjected to 30 minutes of ischemia by transient left anterior descending coronary artery (LAD) ligation. Cardiac function and structure were evaluated 24 hours after reperfusion by echocardiography and histology. Left ventricular ejection fraction was significantly preserved, while infarct size was significantly decreased by CIP4 CKO when compared to the two control cohorts. Separate mice subjected to permanent LAD ligation showed that CIP4 CKO resulted in long-term preservation of cardiac function. To disrupt CaNAβ-CIP4 complexes in vivo, AAV has been used to express the CaNAβ PP domain peptide. In contrast to the expression of the VIVIT pan-CaN inhibitory peptide, CaNAβ PP peptide expression did not worsen I/R injury. Together, these results suggest that PP-anchored CaNAβ does not contribute to cardioprotection. Targeting CIP4 signalosomes may constitute a novel strategy for inhibiting I/R injury and preventing the development of heart failure in response to diverse forms of cardiovascular disease.
Abstract P3119: Identifying The Windows Of Cardiac Risk And Opportunity In Menopause
Authors: Ciara Barry, Sarah Rouhana, Jinan Saboune, Jessica L Braun, Mia S Geromella, Val A Fajardo, W. G Pyle
Journal: Circulation Research (2023)

Post-acute myocardial infarction (AMI) mortality has significant sex differences with a bias towards higher rates in women. While treatment inequities and access to care profoundly impact outcomes, biological factors also contribute to survival discrepancies. One complicating factor that drives worse outcomes in women is menopause. Understanding the biological basis for AMI mortality risk and the exacerbating role of menopause has been hampered by the lack of an animal model that recapitulates the dynamic changes associated with the perimenopausal transition. We used a mouse model of menopause in which ovarian failure is gradually induced following treatment with 4-vinylcyclohexene diepoxide (VCD, 160 mg/kg, 15d), allowing for examination of cardiac alterations established during perimenopause. Echocardiography revealed no contractile changes during perimenopause, but cardiac myofilament activation was significantly affected. Calcium re-uptake by SERCA was impaired by the end of perimenopause. Changes in calcium handling were associated with alterations in calcium handling protein expression and phosphorylation. Activation of RISK and SAFE signaling molecules that protect against ischemia-reperfusion (IR) injury was altered throughout perimenopause, and the RISK-SAFE response to IR differed at points during perimenopause. The myocardial response to cardioprotective estrogen receptor activation also varied throughout perimenopause. Together these data reveal significant, nonlinear alterations in myocardial physiology and endogenous protective signing pathways during perimenopause. Disruptions in endogenous protection and calcium handling are established prior to menopause, providing possible mechanisms for worse post-menopausal AMI outcomes. The altered response to estrogen receptor activation preceding menopause may explain the inconsistent effectiveness of post-menopausal estrogen replacement therapy.
Abstract P2121: Relative Importance Of Reactive Oxygen Species From Reverse Electron Transfer At Mitochondrial Complex I In Reperfusion Injury
Authors: Caio Tabata Fukushima, Ian-Shika Dancil, Nidhi Shah, Paul S Brookes
Journal: Circulation Research (2023)

The metabolite succinate accumulates during tissue ischemia. In early reperfusion, succinate is rapidly metabolized, driving a burst of mitochondrial reactive oxygen species (ROS) generation that triggers cell death. It is widely believed that the source of succinate-driven ROS in early reperfusion is reverse electron transfer (RET) at respiratory complex I (Cx-I). However, evidence for Cx-I RET ROS is largely based on experimental evidence from isolated mitochondria with succinate as the sole substrate. We hypothesized that numerous other conditions present during early reperfusion may impact Cx-I RET ROS. Thus, experimental perturbations were applied to mitochondria to model early reperfusion, with measurement of ROS via the fluorescent probe PHPA. These conditions included: (i) A high concentration of NADH; (ii) A high concentration of lactate; (iii) A mildly acidic pH; (iv) A non-zero concentration of ADP buffered by a creatine / creatine phosphate / creatine kinase system; (v) Presence of purine nucleotide breakdown products; (vi) Presence of calcium. These studies revealed that under early reperfusion-like conditions mitochondrial ROS generation still occurred via Cx-I RET, but at a magnitude only half that previously reported. Furthermore, residual ROS that was insensitive to a specific Cx-I RET inhibitor (S1QEL) was slightly elevated under early-reperfusion like conditions, and this residual ROS was mostly inhibited by S3QEL, a blocker of ROS from respiratory complex III (Cx-III). Together, these data suggest that the relative contribution of Cx-I RET ROS to reperfusion injury may be overestimated, and that of Cx-III ROS underestimated.
Abstract P2106: Transforming Growth Factor β-dependent Regulation Of Cardiomyocyte Maturation
Authors: Rachel A Minerath, Kelly Grimes, Rajesh K Kasam, Michelle Sargent, Allen York, Yasuhide Kuwabara, Anthony Saviola, Christina Alfieri, Kirk Hansen, Katherine E Yutzey, Jeff D Molkentin
Journal: Circulation Research (2023)

Transforming growth factor β (TGFβ) is a secreted growth factor that is sequestered to the ECM as a latent complex and released in response to cardiac stress. Once released, TGFβ is required for conversion of quiescent fibroblasts to myofibroblasts that support cardiac repair and fibrosis through production of ECM components. While these functional features of TGFβ are well known, the fibroblast-independent effector functions of TGFβ in the heart are unknown. We hypothesized that TGFβ is a critical ECM-cell cross talk mediator in the heart whereby cardiomyocyte -generated TGFβ communicates with fibroblasts to program the composition and stability of the ECM in a reinforcing feedback network. We found that deletion of TGFβ ligands ( Tgfb1 , Tgfb2 , and Tgfb3 ) from cardiomyocytes with an α-MHC-Cre transgene ( Tgfb123 fl/fl -αMHC-Cre ), but not a fibroblast-specific targeted allele ( Tgfb123 fl/fl -Tcf21-MCM ), results in cardiac dysfunction. These results suggest that cardiomyocytes (CMs) are the predominant source of TGFβ in the heart at baseline and that perhaps celltypes beyond the fibroblast were also involved. Interestingly, fibroblast numbers and ECM deposition are increased in Tgfb123 fl/fl -αMHC-Cre hearts and with acute injury hearts from these mice produce shorter, disorganized collagen fibers. Gene expression profiling of Tgfb123 fl/fl -αMHC-Cre hearts at 6 weeks of age demonstrates defective cardiomyocyte maturation. Indeed, Tgfb123 fl/fl -αMHC-Cre hearts are dysmorphic at neonatal day 1 and cardiomyocytes from these hearts remain in cell cycle for a longer duration with greater levels of mononucleation. However, deleting TGFβ receptors I/II or Smad2/3 from CMs does not recapitulate this phenotype suggesting that TGFβ is not driving CM maturation in an autocrine manner. Instead, these findings indicate that TGFβ functions as a critical cross-talk mediator to fibroblasts to establish proper ECM organization and content that drives CM maturation.
Abstract P1146: Global Analysis Of Murine Cardiac Gene Program Regulated By Thyroid Hormone And Transcriptional Repressor Rest
Authors: Megan L Russell, Bingruo Wu, Pengfei Lu, Deyou Zheng, Bin Zhou
Journal: Circulation Research (2023)

Background: During adult heart failure, transcriptional reprogramming of fetal cardiac genes coincides with dysregulated heart functions. However, the underlying mechanisms remain incompletely understood. Thyroid hormone and RE1 silencing transcription factor (REST) are known regulators of cardiac gene reprogramming. Here, we report a global analysis of their co-regulatory functions. Methods: We generated a dual Myh6 and Myh7 mouse reporter line. This line was crossed with a conditional Rest allele to generate myocardial Rest knockout mice. Control and Rest null mice were treated with propylthiouracil (PTU) to inhibit thyroid hormone synthesis. PTU treatment induced heart failure and adult-to-fetal cardiac gene remodeling. Single nuclei multiomic profiling was performed, linking gene expression and gene regulatory network information from each nuclei to elucidate the impact of PTU or REST or both on cardiac gene reprogramming. Samples were collected to validate the newly identified factors involved in reprogramming. Results: Data analysis showed that PTU treatment upregulates transcription factor binding involving circadian gene expression ( Cry2 , Per2 , Per3 ). Downregulated genes are related to voltage-gated channel activity and BMP/TGFB activities involving heart development ( Myh6 , Nrg1 , Bmpr1a ) and SA node action potential ( Scn5a , Hcn2 , Ryr2 ). In contrast, Rest knockout upregulated genes are associated with arrhythmia, i.e. , cAMP catabolism ( Pde4d , Pde7a , Pde7b ), heart conduction ( Cacna1c , Gja1 , Pln ), and ion transport ( Kcnip2 , Pde4d, Esr1 ). Downregulated genes are involved in protein functions involving integrin-mediated signaling ( Itgal , Itga4 , Itga5 ), cytoskeleton organization ( Ctnna2 , Actb , Xirp1 ), and fiber organization ( Tgfb3 , Smad3 , Ptk2b ). Furthermore, a subset of genes was found to be co-regulated by thyroid hormone and REST, including Ccn2 , Myh9 , Nmrk2 , Fgr , Itga4 , Prkcb , Cspg4 which are involved in integrin signaling pathway, immune activation, and angiogenesis. Conclusion: Thyroid hormone and REST have shared function in cardiac gene reprogramming involving cardiac conduction and fibrosis as well as their own newly discovered, unique roles regulating circadian genes and cAMP catabolism, respectively.
Abstract P2127: A Novel Arnt Dependent Hif2a Singling In Protecting Against Cardiac Microvascular Endothelial Barrier Dysfunction Following Myocardial Infarction
Authors: Karim Karim Ullah, Lizhuo Ai, Yan Li, Lifeng Liu, Kaichao Pan, Zainab Humayun, Albert Sitikov, Qiaozhu Su, Qiong Zhao, Willard W Sharp, Yun Fang, David Wu, James K Liao, Rongxue R Wu
Journal: Circulation Research (2023)

Rationale: Cardiac microvascular leakage and inflammation are triggered during myocardial infarction (MI) and contribute to heart failure. Hypoxia-inducible factor 2α ( Hif2α ) is highly expressed in endothelial cells (ECs) and rapidly activated by myocardial ischemia, but its impact in microvascular endothelial barrier function during MI is unclear. Objective: To test our hypothesis that the expression of Hif2α in ECs regulates cardiac microvascular permeability in infarcted hearts, which is through its binding partner aryl hydrocarbon nuclear translocator. (ARNT). Methods and Results: Experiments were conducted with mice carrying an inducible EC-specific Hif2α -knockout ( ecHif2α -/- ) mutation, with mouse cardiac microvascular endothelial cells (CMVECs) isolated from the hearts of ecHif2α -/- mice after the mutation was induced, and with human CMVECs and umbilical-vein endothelial cells transfected with ecHif2α siRNA. After MI induction, echocardiographic assessments of cardiac function were significantly lower, while measures of cardiac microvascular leakage (Evans blue assay), plasma IL6 levels, and cardiac neutrophil accumulation and fibrosis (histology) were significantly greater, in ecHif2α -/- mice than in control mice, and RNA-sequencing analysis of heart tissues from both groups indicated that the expression of genes involved in vascular permeability and collagen synthesis was enriched in ecHif2α -/- hearts. In cultured ECs, ec Hif2α deficiency was associated with declines in endothelial barrier function (electrical cell impedance assay) and the reduced abundance of tight-junction proteins, as well as an increase in the expression of inflammatory markers, all of which were largely reversed by the overexpression of ARNT. We also found that ARNT, but not Hif2α, binds directly to the IL6 promoter and suppresses IL6 expression. Conclusions: Endothelial HIF-2a protects from hypoxia-induced cardiac microvascular barrier dysfunction, promotes inflammation damage and represents a potential therapeutic target for cardioprotection, and prevention of fibrosis following acute ischemic injury.